Biologists’ Clever Way to Detect Animals They Can’t Find

Wildlife doesn’t get much weirder than the hellbender, a frilly, crayfish-gobbling salamander, about the length of a baby alligator, whose bizarre aliases include “snot otter,” “devil dog,” and “grampus.” The giant amphibian stalks rocky streambeds throughout the eastern United States—or at least it did, until agriculture, deforestation, and dams ruined water quality and habitats throughout much of its range. These days, scientists aren’t certain where snot otters still roam.

“They like to hide under big rocks, they’re nocturnal, and they blend in perfectly with the stream bottom,” says Andy Adams, a Loyola University biology professor who hasn’t spotted a hellbender since he was himself an undergraduate. “Even people who study them rarely see them.”

The creature’s scarcity and secretive habits have created a dilemma for conservation. How can agencies protect hellbenders when they can’t even be sure where they are? The answer: Environmental DNA, or eDNA, the scientific technique that’s revolutionizing aquatic biology.

For decades, searching for snot otters required scientists to spend long days crawling along streambeds and flipping rocks—a method that was arduous, disruptive to salamanders, and fraught with uncertainty. By contrast, eDNA could hardly be more convenient, for researcher and subject alike: Just take a few quarts of water back to your laboratory and test for traces of your target organism’s DNA, which could come from shed skin, reproductive material, or even feces. No longer must you lay eyes on a live hellbender to know they persist; instead, finding a molecular fingerprint is evidence enough.

This fall, Adams and the Susquehannock Wildlife Society began using eDNA to hunt for hellbenders in Maryland’s Susquehanna River and its tributaries—one small piece of a five-state effort to pin down the cryptic critter’s population status. “We’re trying to figure out not only where hellbenders are present, but also how many there are,” says Kimberly Terrell, wildlife biologist at the Smithsonian Institution, who’s coordinating the extensive study. “How well can we really estimate abundance using this tool?”

Hellbenders are far from the only species to be tracked down with eDNA. The technique has been used since 2008 to detect animals from Italian crested newts to chinook salmon to, most famously, Asian carp, devastating invasive fish that’ve turned some Midwestern rivers into piscine warzones. Scientists have turned to eDNA to track the invader’s advance. “It’s like a smoke detector,” says Lindsay Chadderton, an aquatic biologist at the Nature Conservancy. “It’s our first indication that there’s something there.”

Still, traditional eDNA techniques are fairly blunt instruments. Environmental DNA surveys rely on the polymerase chain reaction (PCR), a way to amplify a few copies of a piece of DNA into a few million. PCR is among the most important methods in molecular biology, so foundational that it won Kary Mullis a Nobel Prize (and inspired at least one truly cheese-tastic song). But when it comes to identifying eDNA, traditional PCR (aka endpoint PCR or ePCR), comes with a drawback: It can’t easily tell you howmuch DNA is floating around. It’s not all that sensitive, either: Some sites known to host Asian carp have tested negative for their genetic material.

That imprecision can lead to controversy. Last year, traces of Asian carp DNA were found in Lake Michigan, a long-dreaded nightmare for state and federal agencies who fear the carp could crowd out natives. But while Chadderton suspects the samples came from live fish, other scientists believe the genetic material could have derived from indirect sources—a plume of bird droppings, maybe, or the hull of a fisherman’s boat. And those aren’t impossible scenarios: One study found that carp DNA stays viable in slime or eagles feces for up to a month.

Fortunately a new alternative to traditional PCR, called quantitative PCR or qPCR, might someday settle matters. qPCR allows scientists to follow the amplification process as it happens, instead of after the cycles are complete; by observing amplification in real time, researchers can obtain invaluable information about the amount of DNA in a sample. “eDNA is inherently indirect—you’re just making an inference about whether an organism was here,” says Cameron Turner, a Notre Dame PhD student who studies the genetic detection of invasive species. “But qPCR at least provides the possibility of a more definitive answer.”

That possibility is tantalizing to researchers like Turner, who earlier this month published a specific qPCR test for Asian carp. When he compared qPCR against traditional PCR, he found the new method was 22 times more likely to notice carp in a pond. An additional methodological tweak allowed Turner to detect 5 times more DNA in the water. The refinements may eventually help resolve questionable cases like the Great Lakes. Says Turner: “Now that we can measure the amount of DNA, we may be able to establish some thresholds that allow us to say, okay, at these levels, there are probably live organisms.”

Andy Adams’ Susqehanna River samples, too, will be processed with qPCR, which has proved effective on hellbenders as well as carp. Adams and his team surveyed three tributaries early this fall, when spawning snot otters fill streams with sperm and eggs. After filtering river water to catch strands of biological material, Adams rolled up the paper filters, stuck them into a vial with a preservative, and shipped them off to a lab at Buffalo State College for analysis. “To know where hellbenders are in this state is going to be huge,” says Adams.

Powerful though it may be, however, even qPCR has its limits. Knowing how many hellbenders dwell in a river, for instance, doesn’t answer important, finer-grain questions about population health or habitat quality. “You might have a degraded, low-quality stream that only has a couple of animals hanging on, or you might have a clean stream with a healthy population that’s naturally low density,” says Kimberly Terrell. “eDNA might not be able to distinguish between those two scenarios.” For now, some questions are still best answered by good old-fashioned rock-flipping.

Ben Goldfarb is a Seattle-based environmental journalist and correspondent at High Country News. He tweets @ben_a_goldfarb.